Tuesday, 30 May 2017

Modulating Neuronal Chloride via WNK

Today’s post is a little complicated,
but should be relevant to parents already using bumetanide to reduce the
severity of autism.

Tuning neurons via Cl-sensitive WNK

The science behind today’s post only
started to evolve twenty years ago when it became understood how chloride enters
and exits the neurons in your brain. Nonetheless there is now a vast amount of
research and there are parts that have not yet been covered in this blog.

A moving target

The first thing to realize is that
trying to reduce the elevated level of chloride found in much autism is very
much an ongoing battle. Chloride is flowing in too fast via NKCC1 and exiting
too slowing via KCC2.

If you want to reduce the entry via
NKCC1, or increase the exit via KCC2, either of these two strategies should
lower the equilibrium level of chloride.Most strategies in this blog target NKCC1, but in another disease
(neuropathic pain) the target has been KCC2.

Whichever you target, the risk is that
the body’s feedback loops come into play and undo some of your good work. This was
highlighted recently in a paper by Kristopher Kahle at Yale, who looks likely to be
joining this blog’s Dean’s List, which highlights the researchers who are
really worth following. He is part of the new generation of higher quality
researcherswho have an interest in autism.

If all that was not complex, we have
to realize that the number of these valves (cotransporters) that either let
chloride enter or exit, is changing all the time.Many factors relating to inflammation and
pain affect the number of NKCC1 and KCC2 cotransporters, so in times of inflammation
you get a reduction KCC2 and/or an
increase in NKCC1; hence a higher level of chloride in your neurons.

When people have a traumatic brain
injury (TBI), they get an increase in NKCC1 and so an increase in neuronal
chloride.This makes the
neurotransmitter GABA less inhibitory, this can lead to cognitive loss,
behavioral changes and even a tendency to seizures.

In TBI not surprisingly you have
elevated inflammatory signaling, such as via something called NF-κB. As pointed out by our reader AJ, when you take the
supplement Astaxanthin, you reduce the expression of NKCC1 in TBI and this has
been shown to be via NF-κB. So the potent antioxidant and broadly
anti-inflammatory Astaxanthin is a good choice for people with elevated NF-κB.

Much is written in
neuropathic pain research about KCC2 and drugs are being developed that could
later be repurposed for autism (and indeed TBI). In neuropathic pain there is a
lack of KCC2 expression and this is known to be linked to something called
WNK1.The WNK1 gene provides
instructions for making multiple versions of the WNK1 protein.

Mechanisms that control NKCC1
and KCC2

There are multiple mechanisms
that affect the expression of NKCC1 and KCC2.In some cases the two (NKCC1 and KCC2) are interrelated so either one is
expressed or the other is expressed.In
the mature brain there should be KCC2, but little NKCC1.

The current research by Kristopher Kahle
is based on the recent discovery
of a “rheostat” of chloride homeostasis, comprising the Cl- sensitive
WNK-SPAK kinases and the NKCC1/KCC2 cotransporters. This rheostat provides a way
to reversibly tune the strength of inhibition in neurons.

In effect this means
that inhibiting WNK should make GABA more inhibitory, which is the goal for all
people who have elevated chloride in their neurons.

GABAA receptors are ligand-gated Cl- channels. GABAAR
activation can elicit excitatory or inhibitory responses, depending on the
intraneuronal Cl- concentration levels. Such levels are largely established by
the Cl- co-transporters NKCC1 and KCC2. A progressive postnatal increase in
KCC2 over NKCC1 activity drives the emergence of GABAAR-mediated
synaptic inhibition, and is critical for functional brain maturation. A delay in this NKCC1/KCC2
‘switch’ contributes to the impairment of GABAergic inhibition observed in Rett
syndrome, fragile X syndrome, and other neurodevelopmental conditions, such as
epilepsy.

Kristopher Kahle and his colleagues aim to understand the
mechanisms that govern these developmental changes in NKCC1/KCC2 activity. They
hypothesize that an improved knowledge of these mechanisms will lead to the
development of novel strategies for restoring GABAergic inhibition. The researchers propose to
exploit their recent discovery of a ‘rheostat’ of Cl- homeostasis, comprising
the Cl-sensitive WNK-SPAK kinases and the NKCC1/KCC2 cotransporters1-3.
This rheostat provides a phosphorylation-dependent way to reversibly tune the
strength of synaptic inhibition in neurons.

The team will create genetic mouse models with inducible
expression of phospho-mimetic or constitutively dephosphorylated WNK-SPAK-KCC2
pathway components. They will also develop novel WNK-SPAK kinase inhibitors
that function as simultaneous NKCC1 inhibitors and KCC2 activators. These mouse
models and compounds will be used to therapeutically restore GABA inhibition in
the Rett syndrome MeCP2(R308/Y) mouse model. The researchers will use a
combination of two-photon microscopy coupled with improved fluorescent
optogenetic Cl- sensing, quantitative phosphoproteomics and patch-clamp
electrophysiology to assess cellular and physiological changes in these mice.

The intracellular concentration of Cl− ([Cl−]i)
in neurons is a highly regulated variable that is established and modulated by
the finely tuned activity of the KCC2 cotransporter. Despite the importance of
KCC2 for neurophysiology and its role in multiple neuropsychiatric diseases,
our knowledge of the transporter's regulatory mechanisms is incomplete. Recent
studies suggest that the phosphorylation state of KCC2 at specific residues in
its cytoplasmic COOH terminus, such as Ser940 and Thr906/Thr1007, encodes
discrete levels of transporter activity that elicit graded changes in neuronal
Cl− extrusion to modulate the strength of synaptic inhibition via Cl−-permeable
GABAA receptors. In
this review, we propose that the functional and physical coupling of KCC2 to Cl−-sensitive
kinase(s), such as the WNK1-SPAK kinase complex, constitutes a molecular
“rheostat” that regulates [Cl−]i and thereby influences
the functional plasticity of GABA. The rapid reversibility of
(de)phosphorylation facilitates regulatory precision, and multisite
phosphorylation allows for the control of KCC2 activity by different inputs via
distinct or partially overlapping upstream signaling cascades that may become
more or less important depending on the physiological context. While this
adaptation mechanism is highly suited to maintaining homeostasis, its
adjustable set points may render it vulnerable to perturbation and
dysregulation. Finally, we
suggest that pharmacological modulation of this kinase-KCC2 rheostat might be a
particularly efficacious strategy to enhance Cl− extrusion and
therapeutically restore GABA inhibition.

Dominant-negative mutation, genetic knockdown, or
chemical inhibition of WNK1 in immature neurons (but not mature neurons) is
sufficient to trigger a hyperpolarizing shift in GABA activity by enhancing
KCC2-mediated Cl− extrusion secondary to a reduction of
Thr906/Thr1007 inhibitory phosphorylation (Friedel et al. 2015). These results extended previous work by Rinehart et al. (2009), who showed that KCC2 Thr906 phosphorylation
inversely correlates with KCC2 activity in the developing mouse brain, and Inoue et al. (2012), who showed a phosphorylation-dependent
inhibitory effect of taurine on KCC2 activity in immature neurons that was
recapitulated by WNK1 overexpression in the absence of taurine. Together, these compelling data
suggest that a postnatal decrease in WNK1-regulated inhibitory phosphorylation
of KCC2 also contributes to increased KCC2 function(Fig. 5),
and thus to the excitatory-to-inhibitory GABA shift that occurs during
development.This
also raises the possibility that dysfunctional phosphoregulation of these sites
could be important in certain neurodevelopmental pathologies, like autism or
neonatal seizures. An
important issue of future investigation will be to determine how the increased
levels of Cl− in immature neurons affect WNK1 kinase activity. Could
taurine, a factor known to activate WNK1 in immature neurons, achieve this by
decreasing the sensitivity of WNK1 to Cl−?

Recently, a few groups have developed innovative
high-throughput assays to screen for compounds that modulate KCC2 activity (Delpire et al. 2009, 2012; Gagnon et al. 2013), and one drug shows promise as a KCC2-dependent Cl− extrusion
enhancer with therapeutic effect in a model of neuropathic pain (Gagnon et al. 2013). These early but encouraging results
require validation, but they establish the validity in vivo of the concept of
GABA modulation via the pharmacological targeting of CCC-dependent Cl−
transport (Gagnon et al. 2013; Kahle et al. 2014a; Kaila et al. 2014). Could CCC phosphoregulatory mechanisms, normally employed to modulate
transporter activity in response to perturbation or biological need, be
harnessed to stimulate the KCCs (or inhibit NKCC1) for therapeutic benefit in
disease states featuring an accumulation of intracellular Cl−?

Moreover, since
the WNK kinases might also be the Cl− sensors that detect changes in
intracellular Cl− (Piala et al. 2014), inhibiting these molecules might prevent
feedback mechanisms that would counter the effects of targeting NKCC1 or KCC2
alone.

The
K(+)-Cl(-) cotransporter KCC2 is responsible for maintaining low Cl(-)
concentration in neurons of the central nervous system (CNS), which is
essential for postsynaptic inhibition through GABA(A) and glycine receptors.
Although no CNS disorders have been associated with KCC2 mutations, loss of
activity of this transporter has emerged as a key mechanism underlying several
neurological and psychiatric disorders, including epilepsy, motor spasticity,
stress, anxiety, schizophrenia, morphine-induced hyperalgesia and chronic pain.
Recent reports indicate that enhancing KCC2 activity may be the favored
therapeutic strategy to restore inhibition and normal function in pathological
conditions involving impaired Cl(-) transport. We designed an assay for
high-throughput screening that led to the identification of KCC2 activators
that reduce intracellular chloride concentration ([Cl(-)]i). Optimization of a first-in-class
arylmethylidine family of compounds resulted in a KCC2-selective analog
(CLP257) that lowers [Cl(-)]i. CLP257 restored impaired Cl(-) transport in
neurons with diminished KCC2 activity. The compound rescued KCC2 plasma
membrane expression, renormalized stimulus-evoked responses in spinal
nociceptive pathways sensitized after nerve injury and alleviated
hypersensitivity in a rat model of neuropathic pain. Oral efficacy for
analgesia equivalent to that of pregabalin but without motor impairment was
achievable with a CLP257 prodrug. These results validate KCC2 as a drugable target for CNS diseases.

WNK1 [with no
lysine (K)] is a serine-threonine kinase associated with a form of familial
hypertension. WNK1 is at the top of a kinase cascade leading to phosphorylation
of several cotransporters, in particular those transporting sodium, potassium,
and chloride (NKCC), sodium and chloride (NCC), and potassium and chloride
(KCC). The
responsiveness of NKCC, NCC, and KCC to changes in extracellular chloride
parallels their phosphorylation state, provoking the proposal that these
transporters are controlled by a chloride-sensitive protein kinase. Here, we
found that chloride stabilizes the inactive conformation of WNK1, preventing
kinase autophosphorylation and activation. Crystallographic studies of inactive
WNK1 in the presence of chloride revealed that chloride binds directly to the
catalytic site, providing a basis for the unique position of the catalytic
lysine. Mutagenesis of the chloride binding site rendered the kinase less sensitive
to inhibition of autophosphorylation by chloride, validating the binding site. Thus, these data suggest that
WNK1 functions as a chloride sensor through direct binding of a regulatory
chloride ion to the active site, which inhibits autophosphorylation.

The WNK-SPAK/OSR1 kinase complex is composed of the
kinases WNK (with no lysine) and SPAK (SPS1-related proline/alanine-rich
kinase) or the SPAK homolog OSR1 (oxidative stress–responsive kinase 1). The WNK family senses changes in intracellular Cl− concentration,
extracellular osmolarity, and cell volume and transduces this information to
sodium (Na+), potassium (K+), and chloride (Cl−)
cotransporters [collectively referred to as CCCs (cation-chloride
cotransporters)] and ion channels to maintain cellular and organismal
homeostasis and affect cellular morphology and behavior. Several genes encoding
proteins in this pathway are mutated in human disease, and the cotransporters
are targets of commonly used drugs. WNKs stimulate the kinases SPAK and OSR1, which directly phosphorylate
and stimulate Cl−-importing, Na+-driven CCCs or inhibit
the Cl−-extruding, K+-driven CCCs. These
coordinated and reciprocal actions on the CCCs are triggered by an interaction
between RFXV/I motifs within the WNKs and CCCs and a conserved
carboxyl-terminal docking domain in SPAK and OSR1. This interaction site represents a potentially
druggable node that could be more effective than targeting the cotransporters
directly. In the kidney, WNK-SPAK/OSR1 inhibition decreases epithelial
NaCl reabsorption and K+ secretion to lower blood pressure while
maintaining serum K+. In neurons, WNK-SPAK/OSR1 inhibition could facilitate Cl−
extrusion and promote γ-aminobutyric acidergic (GABAergic) inhibition.Such drugs could have efficacy
as K+-sparing blood pressure–lowering agents in essential
hypertension, nonaddictive analgesics in neuropathic pain, and promoters of GABAergic
inhibition in diseases associated with neuronal hyperactivity, such as
epilepsy, spasticity, neuropathic pain, schizophrenia, and autism.

The
Ste20 family protein kinases oxidative stress-responsive 1 (OSR1) and the
STE20/SPS1-related proline-, alanine-rich kinase directly regulate the solute
carrier 12 family of cation-chloride cotransporters and thereby modulate a
range of processes including cell volume homeostasis, blood pressure, hearing,
and kidney function. OSR1 andSTE20/SPS1-related proline-,alanine-rich kinase
are activated by with no lysine [K] protein kinases that phosphorylate the
essential activation loop regulatory site on these kinases. We found that
inhibition of phosphoinositide 3-kinase (PI3K) reduced OSR1 activation by
osmotic stress. Inhibition of the PI3K target pathway, the mammalian target of
rapamycin complex 2 (mTORC2), by depletion of Sin1, one of its components,
decreased activation of OSR1 by sorbitol and reduced activity of the OSR1
substrate, the sodium, potassium, two chloride cotransporter, in HeLa cells.
OSR1 activity was also reduced with a pharmacological inhibitor of mTOR.
mTORC2phosphorylated OSR1 on S339 in vitro, and mutation of this residue eliminated
OSR1 phosphorylation by mTORC2. Thus, we identify a previously unrecognized
connection ofthePI3K pathwaythroughmTORC2 to a Ste20 proteinkinase and ion
homeostasis.

Signiﬁcance

With no lysine [K] (WNK) protein kinases are
sensitive to changes in osmotic stress. Through the downstream protein kinases
oxidative stress-responsive 1 (OSR1) and STE20/SPS1related proline-,
alanine-rich kinase, WNKs regulate a family of ion cotransporters and thereby
modulate a range of processes including cell volume homeostasis, blood
pressure, hearing, and kidney function. We found that a major phosphoinositide 3-kinase target
pathway, the mammalian target of rapamycin complex 2, also phosphorylates OSR1,
coordinating with WNK1 to enhance OSR1 and ion cotransporter function.

Changes in tonicity regulate the
WNK-OSR1/SPAK pathway to control ion cotransporters for volume and ion
homeostasis. We ﬁnd that mTORC2 also contributes to enhanced OSR1 activity.
Inhibiting mTORC2 does not inhibit WNK1 activity, indicating PF1 and PF2regions.

These data demonstrate that the
WNK-regulated SPAK/OSR1 kinases directly phosphorylate the N[K]CCs and KCCs,
promoting their stimulation and inhibition respectively. Given these
reciprocal actions with anticipated net effects of increasing Cl− inﬂux, we propose that the targeting of
WNK–SPAK/OSR1 with kinase inhibitors might be a novel potent strategy to
enhance cellular Cl− extrusion, with potential implications for the therapeutic
modulation of epithelial and neuronal ion transport in human disease states.

WNK
Inhibitors

The first orally bioavailable
pan-WNK-kinase inhibitor is WNK463.

“WNK463 is an orally bioavailable
pan-WNK-kinase inhibitor. In vivo: WNK463, that exploits unique structural
features of the WNK kinases for both affinity and kinase selectivity. In rodent
models of hypertension, WNK463 affects blood pressure and body fluid and
electro-lyte homeostasis, consistent with WNK-kinase-associated physiology and
pathophysiology.”\

WNK463
is available as a research drug.

It
looks like WNK2 is also very relevant, perhaps more so than WNK1, because we
are interested specifically in the brain, where there is a lot of WNK2. WNK3
also looks very relevant. There is also WNK4.

Here,
we show that WNK2,
unlike other WNKs, is not expressed in kidney; rather, it is a neuron-enriched
kinase primarily expressed in neocortical pyramidal cells, thalamic relay
cells, and cerebellar granule and Purkinje cells in both the developing and
adult brain. Bumetanide-sensitive and Cl−-dependent 86Rb+
uptake assays in Xenopus laevis oocytes revealed that WNK2 promotes Cl−
accumulation by reciprocally activating NKCC1 and inhibiting KCC2 in a
kinase-dependent manner, effectively bypassing normal tonicity requirements for
cotransporter regulation.

WNK3 KO mice exhibited
significantly decreased infarct volume and axonal demyelination, less cerebral
edema, and accelerated neurobehavioral recovery compared to WNK3 WT mice
subjected to MCA occlusion. The
neuroprotective phenotypes conferred by WNK3 KO were associated with a decrease
in stimulatory hyper-phosphorylations of the SPAK/OSR1 catalytic T-loop and of
NKCC1 stimulatory sites Thr203/Thr207/Thr212,
as well as with decreased cell surface expression of NKCC1. Genetic
inhibition of WNK3 or siRNA knockdown of SPAK/OSR1 increased the tolerance of
cultured primary neurons and oligodendrocytes to in vitro ischemia.

CONCLUSION

These data identify a
novel role for the WNK3-SPAK/OSR1-NKCC1 signaling pathway in ischemic
neuroglial injury, and suggest the WNK3-SPAK/OSR1 kinase pathway as a
therapeutic target for neuroprotection following ischemic stroke.

Conclusion

I think we can simplify all of this into:-

We already know that many people with autism benefit from making GABA more
inhibitory.

·Reducing the inflow via NKCC1 using
bumetanide and in future years using drugs which better pass the blood brain
barrier, e.g. the research drug BUM5. Consider improving the potency of the current drug bumetanide using an OAT3 inhibitor that will increase its concentration and half-life, apparently already possible with acetazolamide.

·Increasing the outflow via KCC2, possible
with the research drug CLP257

·Reducing the inflow via AE3, possible with Diamox/acetazolamide

·Substituting Br- for Cl-,
using potassium bromide

·Changing the relative expression of
NKCC2/KCC1

Changing
the relative expression of NKCC1/KCC2

·This can be done today by treating any underlying
inflammation.Inflammation shifts the NKCC2/KCC1
balance in a way that makes GABA more excitatory, which is bad. This might be achieved by targeting IL-6, NF-κB or just treating any GI problems and allergies. Always treat the comorbidities of autism.

·Using WNK inhibitors it will hopefully be possible to
manually tune the NKCC1/KCC2 balance, just like tuning a piano. One pan-WNK-kinase inhibitor is WNK463.

·I continue to believe thatRORα could be an effective way to increase KCC2
expression and this is something that is not so hard to test.

I will be
keeping a look out for further papers by Dr Kahle and be interested in any WNK-SPAK/OSR1 inhibitors
he proposes.If I was him I would start
with WNK463.

There
is more to the story, because naturally I want to see how estradiol relates to
WNK and finally wrap up this subject. Then we will know how to treat the immature neurons often found in autism. A case of forever young.

In a following post I intend to do that;
here is a sneak, but complex, preview.

Hi Peter,My 8yo son has been taking Bumetanide for 3 months at dose 0.5mg 2x/day. Nothing happened for the first 3 weeks or so. Then at about 1 month I saw very slight social gains, which can be contributed to other reasons. These very slight gains continued into the second month. My wife kept insisting to stop Bumetanide because she didn't notice anything. After the second month, I added Creatine, Quercetin, and Flaxseed Oil staggered by 1 week. When I added Creatine, I saw immediate gains in awareness. When I added Quercetine 1 week later, I saw the same immediate improvement in awareness. I haven't noticed anything from Flaxseed Oil, but it doesn't hurt. Now at 3 months, the social gains are impressive. But, I am not sure what caused them. This is the drawback of trying too many things at once. Is it possible that Bumetanide has a slow start and then suddenly gives impressive improvements at 3 month mark? I am afraid to stop anything because I haven't seen my son being social like this ever.

V, anything is possible. If you want to know if bumetanide is helping, stop taking it and in between 2-5 days you will gradually lose any potential benefit. Keep the other supplements unchanged. If you and your wife agree that benefits are indeed lost, then restart bumetanide.

Quercetin has some strong anti-inflammatory effects, which affect the way bumetanide works. My son's pollen allergy reduces the effect of bumetanide.

It speaks to the role of peripheral IL-6 receptors, their impact on the gut microbiome, and the resulting impact of the altered gut microbiome on the brain.

If anyone has any insights, please share. To me, at first blush, IL-6 receptor antagonists appear to be an interesting option for ASD given the potential impact of problematic gut microbiome signaling on the brain through the gut-brain axis.

Of course, it's late (almost 1am for me), so I'm probably not going to get to it until tomorrow (or later today, to be technical), but I'm going to look for natural IL-6 antagonists. The ideal therapeutic would be an anti-IL-6 MAb but we're not going to get easy access to one so natural products / easy to access small molecule meds will be the way to go.

The following article gives a list of natural IL-6 inhibitors: https://selfhacked.com/2014/10/20/interleukin-6/#TopWays_to_Inhibit_IL-6

My son has a homozygous mutation in the LOC541472 gene (rs1800795: GG), which creates more IL-6 than normal. In addition, he has allery to pollen, dust, etc. I have seen great improvements in my son from Quercetin, which reduces IL-6, but also stabilizes mast cells & reduces CRH hormone that stresses adrenal glands. EGCG (extract from green tea) is next on my list to try. Like Quercetin, it reduces IL-6, stabilizes mast cells. Interestingly, there are clinical studies of EGCG for Down syndrome. Apparently, it inhibits enzyme DYRK1A, which is over expressed in Down syndrome (due to duplication or triplication of the corresponding gene) and which is the main genetic marker of the disease, responsible for facial features and low cognition.V

Peter, have you seen this very recent paper on modulating E-I imbalance in adults with autism. They did that by modulating GABA/glutamate balance in prefrontal cortex with riluzole.

Not only did riluzole increase "the minimal/absent baseline prefrontal functional connectivity in the ASD group towards the control levels; and there was no longer a group difference in functional connectivity post riluzole”.

Even more interestingly is the following observation (remember the pararoxical benzo effect in some!), my caps:

“Our results indicate that despite a comparable baseline, the E–I response to a riluzole challenge was DIAMETRICALLY OPPOSITE in men with ASD and controls. In controls, the decrease in the inhibitory index (or GABA fraction) was correlated with an increase in glutamate–glutamine (Glx). In ASD, riluzole increased the inhibitory index (or GABA fraction) without changing Glx.

Overall, this pattern of results indicates that riluzole shifts flux towards GABA in ASD and towards glutamate–glutamine in controls.

This unusual E–I responsivity in ASD may help explain other paradoxical findings from studies of the way people with ASD respond to E–I acting medications. For example, GABAA/benzo-diazepine receptor agonists typically have an inhibitory effect in non-ASD populations, but can sometimes cause excitation in individuals with ASD. This opposite direction of response in ASD is important because the initial selection of drug candidates for clinical trials in ASD is often based on their action in unaffected individuals. If, however, the nature of the brain response in ASD fundamentally differs from other populations then similar out-comes cannot be anticipated, and trials are likely to be unsuccessful."

https://www.ncbi.nlm.nih.gov/pubmed/28534874

PS ‘even in adults’ :)

"In conclusion, using [1H]MRS and fMRI, we found that E–I flux and functional connectivity of the prefrontal cortex are differentially regulated in adults with ASD compared with the controls. Importantly, inhibitory tone and functional connectivity can be shifted pharmacologically —and even in adults with ASD."

I wonder how much Riluzole is different from Lamictal. Anyone has any ideas? On the surface, both seem to reduce glutamate release from presynaptic terminals. My son is taking Lamictal for his abnormal EEG. It helped with socialization.V

Riluzole does affect some sodium channels, but it also upregulates EAAT2/GLT-1, which is the principal transporter that clears the excitatory neurotransmitter glutamate from the extracellular space at synapses. This effect is shared by many common antibiotics and many people say their autism improves while on (beta-lactam) antibiotics.

It looks like in some people with autism part of the problem is that glutamate hangs around too long after it has been released and it just build up. It was not a problem of too much released, just a problem with clearance afterwards.

The general finding in the research was that people who have more opioid receptors, tend to laugh more in a social setting. They also found that laughter induced endorphin release from the caudate nucleus, thalamus, and insula (all areas that seem to pop up as problem areas in autism research).

More specifically, off the top of my head I have read multiple studies suggesting an enlarged caudate in autism, a hyperactive thalamus that does not filter sensory information properly, as well as a hyperactive salience network whose principle nodes are the anterior insula and dorsal anterior cingulate cortex.

Now my son will sometimes go on laughing binges that are incomprehensible and when this happens he gets very hyper and then starts doing laps around the house and laugh like crazy. One could just write this off as him being happy, but this behavior is often sensory driven by videos of people acting silly and/or laughing.

So the idea here is that if your autistic child is laughing excessively to some sort of stimming media to the point they get so excited they engage in SIB, it might be a sign of excessive mu-opioid signaling via excess endorphin release or else excessive mu-opiod receptors (i.e. hypersensitivity to opioids) and blockling some of those receptors with naloxone/naltrexone or aspartic acid (what has worked for me since the other two options are not available to me), may help in other areas such as attention and cognition.

We have also trialed butenamide at 0.5 mg twice daily, after a month I didn't notice too much difference, I bumped up to 1 mg twice daily and we are noticing a difference after about a month on that dose. Much more responsive to commands, calmer, less vocal protests during ABA, NO throwing, if in room with 3 people talking I said "close the door" and did this - before would not even register I was requesting anything. We are still hoping for language, butenamide has not improved this in fact over time we have seen less words, maybe 1-2 per day only when prompted with something she wants. I am not sure if that is effect of butenamide or not, with the other gains we will keep going with butenamide for now and keep working on language. She is 5. Thank you Peter, it is through this blog I found out about butenamide and so many other ideas to try. If only she would take everything I need to give her on a daily basis! She will not try NAC in any way I give it. Or magnesium to reduce glutamate. I have tried a million ways. I'll keep trying.

I have a son who is five plus and last year we trialled NAC as well as bumetanide, 1mg once daily. NAC, 600 mg twice and thrice daily, had subtle but clear effects and I attribute it to NACs effect on glutamate. The hugest effect was on his ability to ignore his penile erections, which he is very sensitive to and which he gets under conditions of relaxation, before sleep as well as stimulation, say while doing an assignment. But it started causing digestive issues in my son. Bumetanide seemed to be of no use but after around three weeks on it, my son started bidding goodbye to people, unprompted. Both the interventions affected his motor skills positively, handwriting being one, which is the biggest and earliest indicator of something being effective on him. However, managing the side effects of bunetanide were a bit problematic for me and we did notice a negative effet on speech, both in terms of usage abd clarity, which I sm not sure what to attribute to after one month of bumetanide use. Planning to retrial soon.

The positive effects of bumetanide will likely develop, with an increased awareness of what is going on around her. This will make your behavioral therapy more effective and over time should help with speech, but it all develops gradually.

There are many other antioxidants and they should all help to some extent.

I forgot to mention that once up to 1 mg twice daily, my daughter started going on the potty spontaneously. We have been trying to potty train with ABA for months, but this was new - she ran to the toilet sat on it and did her business all on her own, no prompts, no reinforcers. This effect has continued when at home and I keep the diaper off. In regards to antioxidants, I am doing coQ10 and PPQ, high dose folinic (20 mg) even though she was negative for anti-folate antibodies, vitamin c and some vitamin E. Mostly chewable as she can't swallow capsules. If I can find chewable astaxanthin i will add that next. The language issue is hard as when I was transitioning from 0.5 mg to 1 mg I took 3 days off in those 3 days I did notice maybe a few extra words per day, which reduced again when I gave her the 1mg bid. I wonder of the mechanism. The cognitive gains are there though and your theory that communication will come if cognition is there makes sense to me so we are sticking to it.

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About this Blog

This blog is mainly a review of the science behind autism, looking for pointers to effective treatments for classic autism.The first treatment, Bumetanide, I stumbled upon before starting this blog.The last treatment, tiny doses of Clonazepam, came from a recent paper, highlighted by a regular follower of this blog.You do need some basic scientific knowledge, but putting our minds together, we can make our own medical advances; so all comments and case histories are very welcome.

If your interest is regressive autism, very likely the cause is mitochondrial disease. Classic autism therapies may well be ineffective. Mitochondrial disease can be diagnosed and treated.

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About Me

I am an independent researcher, trawling through the scientific literature and doing some experiments along the way. I am not a doctor. I do have a Master's degree from a top science-only university and another one from a top business school. More relevant is my motivation.

I am developing a novel drug therapy, the Autism Polypill, to treat classic early-onset autism, since this is the type that affects my son. His type of autism is characterized by autistic behaviours, pollen allergy, asthma, some SIB, high serotonin, high cholesterol, high euthyroid, high IGF-1, but without seizures, GI problems, food intolerance or severe MR. I think that ADHD and Asperger's are likely to be very mild forms of this phenotype of autism. Many other types of "autism" are entirely different. As this blog shows, at least classic autism is treatable using today's drugs.